Sequential Actions of the Two Component Peptides of the Lantibiotic Lacticin 3147 Explain Its Antimicrobial Activity at Nanomolar Concentrations

Morgan, Sheila; O’Connor, Paula M.; Cotter, Paul D.; Ross, R. Paul; Hill, Colin

doi:10.1128/aac.49.7.2606-2611.2005

Cited by 107 publications

(92 citation statements)

References 33 publications

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“…In this way, the following pORI280-ltnA1A2 derivatives were made: pOri-ltnA1S7T (primer pair [21][22], pOri-ltnA1S7G (primer pair 24-25), pOri-ltnA1S7V (primer pair 27-28), pOri-ltnA2S9T (primer pair 30-31), pOri-ltnA2S9G (primer pair 33-34), pOri-ltnA2S9V (primer pair 36-37), pOriltnA2S9,12T (primer pair 39-40), pOri-ltnA2S9,12G (primer pair 42-43), and pOri-ltnA2S9,12V (primer pair 45-46). Screening of candidate EC101 transformants was facilitated by the use of check primers 23,26,29,32,35,38,41,44, and 47, respectively, in conjunction with either primer 7 or 8, depending on the orientation of the check primer. In all cases successful mutagenesis was confirmed by DNA sequencing.…”

Section: Methodsmentioning

confidence: 99%

“…This result is almost certainly caused by the isolation procedure, which relies on desorption of the peptides, presumably from a receptor͞ immunity protein on the surface of the producer cell. Indeed, we have previously shown that binding to target cells is a two-step process mediated by LtnA1 (32). Consequently, in the absence of LtnA1, LtnA2 is likely to be lost in the supernatant because it has no receptor peptide to facilitate binding.…”

Section: Identification Of a Dha Reductase Ltnj Responsible For The Smentioning

confidence: 99%

See 1 more Smart Citation

Posttranslational conversion of l -serines to d -alanines is vital for optimal production and activity of the lantibiotic lacticin 3147

Cotter

O’Connor²,

Draper³

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

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120

146

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As a general rule, ribosomally synthesized polypeptides contain amino acids only in the L-isoform in an order dictated by the coding DNA͞RNA. Two of a total of only four examples of L to D conversions in prokaryotic systems occur in posttranslationally modified antimicrobial peptides called lantibiotics. In both examples (lactocin S and lacticin 3147), ribosomally encoded L-serines are enzymatically converted to D-alanines, giving rise to an apparent mistranslation of serine codons to alanine residues. It has been suggested that this conversion results from a two-step reaction initiated by a lantibiotic synthetase converting the gene-encoded L-serine to dehydroalanine (dha). By using lacticin 3147 as a model system, we report the identification of an enzyme, LtnJ, that is responsible for the conversion of dha to D-alanine. Deletion of this enzyme results in the residues remaining as dha intermediates, leading to a dramatic reduction in the antimicrobial activity of the producing strain. The importance of the chirality of the three D-alanines present in lacticin 3147 was confirmed when these residues were systematically substituted by L-alanines. In addition, substitution with L-threonine (ultimately modified to dehydrobutyrine), glycine, or L-valine also resulted in diminished peptide production and͞or relative activity, the extent of which depended on the chirality of the newly incorporated amino acid(s).antimicrobial ͉ bacteriocin ͉ chirality T he presence of D-amino acids in ribosomally synthesized peptides was first observed in dermorphins, peptides found in the skin secretions of species of a subfamily of South American tree frogs, Phyllomedusinae. Subsequently, D-amino acids have been identified in a number of other ribosomally synthesized peptides of eukaryotic origin (1, 2). -Agatoxin IVb and bombinin H, produced by the funnel-web spider and the skin secretions of the frog Bombina variegata, respectively, represent the only instances in which the responsible enzymes, both peptide epimerases, have been identified (1, 2). For -agatoxin IVb the relevant L-serine to D-serine conversion is thought to involve a deprotonation͞reprotonation reaction (3). The presence of D-amino acids in many of these peptides is crucial, as demonstrated by the creation of synthetic analogues with Lrather than D-amino acids at the relevant locations, which are devoid of activity (4). The artificial incorporation of D-amino acids can also be beneficial. For example, a growing number of synthetic peptides͞peptide libraries containing D-amino acids have been assembled to capitalize on the ability of such residues to provide improved protease stability [enhanced pharmokinetic profile (5-7)], alter tertiary structure (new structural elements can be assembled that cannot be built solely from L-amino acids), and affect the activity of bioactive peptides [enhanced antibacterial activity with concomitant reduction in cell cytotoxicity (5, 6, 8-11)].In prokaryotic systems, there are only a few examples of L-to D-amino acid conversions. It...

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Section: Methodsmentioning

confidence: 99%

Section: Identification Of a Dha Reductase Ltnj Responsible For The Smentioning

confidence: 99%

Posttranslational conversion of l -serines to d -alanines is vital for optimal production and activity of the lantibiotic lacticin 3147

Cotter

O’Connor²,

Draper³

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

120

146

View full text Add to dashboard Cite

show abstract

“…A recent study on the mode of action of lacticin 3147 demonstrated that Ltn␣ binds to the peptidoglycan precursor lipid II (12), a result that was anticipated because of the structural similarity between Ltn␣ and mersacidin, which also binds to lipid II (13). However, in order for lacticin 3147 to inhibit cell wall biosynthesis and form small pores in the cell membrane, Ltn␤ is also necessary, leading to a proposed model in which the lipid II:Ltn␣ complex recruits Ltn␤ to form an active complex (12,14).…”

mentioning

confidence: 99%

“…This understanding could be used to develop novel lantibiotics based on designing peptide sequences that can be site-specifically modified to yield new products. Given the synergy observed among twocomponent lantibiotics, which display similar or higher activity than the best single-component lantibiotic nisin (14), the possibility of engineering new lantibiotics with therapeutic potential is even greater for these systems.…”

mentioning

confidence: 99%

Discovery and in vitro biosynthesis of haloduracin, a two-component lantibiotic

McClerren¹,

Cooper

Quan

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

235

321

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Lantibiotics are ribosomally synthesized peptides that undergo posttranslational modifications to their mature, antimicrobial form. They are characterized by the unique amino acids lanthionine and methyllanthionine, introduced by means of dehydration of Ser͞Thr residues followed by reaction of the resulting dehydro amino acids with cysteines to form thioether linkages. Two-component lantibiotics use two peptides that are each posttranslationally modified to yield two functionally distinct products that act in synergy to provide bactericidal activity. By using genetic data instead of isolation, a two-component lantibiotic, haloduracin, was identified in the genome of the Gram-positive alkaliphilic bacterium Bacillus halodurans C-125. We show that heterologously expressed and purified precursor peptides HalA1 and HalA2 are processed by the purified modification enzymes HalM1 and HalM2 in an in vitro reconstitution of the biosynthesis of a two-component lantibiotic. The activity of each HalM enzyme is substratespecific, and the assay products exhibit antimicrobial activity after removal of their leader sequences at an engineered Factor Xa cleavage site, indicating that correct thioether formation has occurred. Haloduracin's biological activity depends on the presence of both modified peptides. The structures of the two mature haloduracin peptides Hal␣ and Hal␤ were investigated, indicating that they have similarities as well as some distinct differences compared with other two-component lantibiotics.lanthionine ͉ dehydroalanine ͉ antibiotic

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“…The 2-component lantibiotics (lanthionine and methyllanthionine) are small microbial peptide bacteriocins produced by Gram positive bacteria such as Lactococcus lactis (Lawton et al, 2007). At nanomolar concentration, these peptides actively inhibit multidrug resistant pathogens by targeting the lipid II component of the bacterial cell wall (Morgan et al, 2005). Lactobacilli are also known to produce non-lanthione containing bacteriocins.…”

Section: Production Of Inhibitory Substancesmentioning

confidence: 99%

The Role of Diet, Prebiotic and Probiotic in the Development and Management of Inflammatory Bowel Diseases (IBD)

Abdulamir¹,

Zaman²,

Hafidh³

et al. 2012

Inflammatory Bowel Disease - Advances in Pathogenesis and Management

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Sequential Actions of the Two Component Peptides of the Lantibiotic Lacticin 3147 Explain Its Antimicrobial Activity at Nanomolar Concentrations

Cited by 107 publications

References 33 publications

Posttranslational conversion of l -serines to d -alanines is vital for optimal production and activity of the lantibiotic lacticin 3147

Posttranslational conversion of l -serines to d -alanines is vital for optimal production and activity of the lantibiotic lacticin 3147

Discovery and in vitro biosynthesis of haloduracin, a two-component lantibiotic

The Role of Diet, Prebiotic and Probiotic in the Development and Management of Inflammatory Bowel Diseases (IBD)

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